Wulff-type boronic acid-based glucose sensor

Abstract

The present invention relates to a biosensor for measuring glucose concentration and its use in glucose sensing, wherein the biosensor is given by formula (I): The present invention contains a polymer that includes the portion of JPEG2026527809000044.jpg6164. The biosensor of the present invention is particularly useful in glucose monitoring performed on subjects under intensive care and in situations where glucose monitoring is performed on unconscious subjects.

Description

Related applications

[0001] This application claims priority under European Patent Application No. EP23187710.1, filed on 25 July 2023, which is incorporated herein by reference in its entirety. [Technical Field]

[0002] This invention relates to a biosensor for measuring glucose concentration and its use in glucose sensing. The biosensor of the present invention comprises a polymer (particularly a high molecular hydrogel) containing a boronic acid-based glucose-binding moiety of formula (I). The biosensor of the present invention is particularly useful in situations where glucose monitoring is performed on subjects in intensive care or on unconscious subjects. Accordingly, this invention relates to the development of an osmotic glucose sensor based on osmotic measurement, incorporating a boronic acid glucose-binding moiety (GBM) of formula (I) to obtain a glucose-responsive material with outstanding sensing properties. [Background technology]

[0003] In addition to glucose-sensing applications, polymers capable of delivering active drugs in a glucose-concentration-dependent manner are sought, particularly for the treatment of diseases characterized by pathological glucose concentrations. For example, diabetes mellitus is a glucose dysregulation characterized by the accumulation of glucose in the blood. This breakdown in glucose regulation can result from the pancreatic islets being unable to secrete insulin or from the body being unable to use insulin properly. In the case of type 1 diabetes, the usual treatment consists of multiple daily blood glucose control and subcutaneous injections. However, better blood glucose control can be achieved if the insulin dose can be constantly adapted to blood glucose levels to avoid blood glucose levels that are lower or higher than the normal range and cause harmful complications. For these reasons, closed-loop delivery of insulin is highly desirable. This closed-loop delivery can be achieved by using a glucose-sensing polymer release system that carries insulin.

[0004] However, the use of glucose sensors based on polymers containing glucose-binding moieties (GBMs) may be limited due to the limited sensitivity of such measurements. Furthermore, since the pKa of boronic acids is typically in the range of 7–9, such polymer sensors have been shown to be highly pH-dependent (Skjaervold et al ANESTHESIOLOGY 2011;114:18-20, Worsley et al. J Diabetes Sci Technol Vol 2, Issue 2, March 2008, Strasma et al. DOI:10.1177 / 1932296815585872).

[0005] A paper by Crane, BC et al., published in the Journal of Diabetes Science and Technology, titled "The Development of a Continuous Intravascular Glucose Monitoring Sensor" (https: / / doi.org / 10.1177 / 1932296815587937), discloses a fluorescent hydrogel for glucose sensing containing Wulff-type boronic acid. This document discloses the use of Wulff-type boronic acid in glucose sensing and discusses the stoichiometry of the bond between boronic acid and glucose. This disclosure differs from the present invention, in particular, in that the sensor is based on fluorescence measurement, not on a change in the volume of the hydrogel.

[0006] The paper “A Wulff-type boronate for boronate affinity capture of cis-diol compounds at medium acidic pH condition” (Li et al., Chem.Commun., 2011, 47, 8169-8171) discloses a boronic acid affinity column containing a boronic acid moiety for cis-diol capture. However, this paper does not disclose glucose sensing using the provided polymer.

[0007] Kim et al. ("Polymeric Monosaccharide Receptors Responsive at Neutral pH") describe the synthesis of styrene ADAM monomer and corresponding linear polymers using RAFT. Glucose sensitivity is determined by the change in turbidity of the polymer solution (without hydrogel) (DOI:10.1021 / ja905652w).

[0008] Document US2016 / 109370 discloses a sensor containing a polymer structure (HEAA-BIS) containing 5-acrylamido-2-((dimethylamino)methyl)phenylboronic acid units.

[0009] Document US2008 / 214912 discloses a glucose detection biosensor containing 4-aminomethyl-2-N,N'-dimethylaminomethylphenylboronic acid units in its polymer structure.

[0010] Document CN 102 219 800 B discloses 4-amino-2-(dimethylaminomethyl)phenylboronic acid incorporated into glycidyl methacrylate and polyethylene glycol diacrylate by copolymerization. [Prior art documents] [Patent Documents]

[0011] [Patent Document 1] U.S. Patent Publication No. 2016 / 109370 [Patent Document 2] U.S. Patent Publication No. 2008 / 214912 [Patent Document 3] China Patent No. 102 219 800 [Non-patent literature]

[0012] [Non-Patent Document 1] Skjaervold et al ANESTHESIOLOGY 2011; 114:18-20 [Non-Patent Document 2] Worsley et al. J Diabetes Sci Technol Vol 2, Issue 2, March 2008 [Non-Patent Document 3] Strasma et al. DOI: 10.1177 / 1932296815585872 [Non-Patent Document 4] Crane, BC et al., Journal of Diabetes Science and Technology (https: / / doi.org / 10.1177 / 1932296815587937) [Non-Patent Document 5] Li et al., Chem. Commun., 2011,47, 8169-8171 [Non-Patent Document 6] Kim et al. (DOI: 10.1021 / ja905652w) [Overview of the Initiative] [Problems that the invention aims to solve]

[0013] This invention addresses the challenge of providing a glucose sensor with improved properties. In particular, the biosensor of this invention overcomes the significant pH interference problem found in state-of-the-art biosensors based on boronic acids. Furthermore, in a special embodiment of the invention, the biosensor of this invention features improved selectivity for glucose over fructose and / or mannitol interference, as well as suppression of other common interfering substances. [Means for solving the problem]

[0014] Therefore, the present invention provides a polymer sensor, particularly a polymer hydrogel sensor, that shrinks linearly when exposed to increasing glucose concentrations. It is believed that the presence of an orthoamino group, such as in part (I), lowers the pKa of the boronic acid portion, enabling glucose complex formation at physiological pH (i.e., pH 7.35-7.45) with very low pH dependence within the physiological range.

[0015] Accordingly, this problem is solved as described in the embodiments disclosed herein and as characterized by the claims. The present invention can be summarized in the following aspects.

[0016] In a first embodiment, the present invention relates to a biosensor for measuring glucose concentration, wherein the biosensor comprises formula (I):

[0017] [ka]

[0018] A polymer containing a portion thereof, Here, the portion of formula (I) is fixed to the polymer, In the above formula, each R is independently C 1-5 Alkyl, C 2-5 Alkenyl or C 2-5 It is alkinyl; In the above formula, each R S Independently, C1-5 Alkyl, C 2-5 Alkenyl, C 2-5 Alkynyl, -(C 0-3 Alkylene)-OH, -(C 0-3 Alkylene)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-O(C 1-5 Alkylene)-OH, -(C 0-3 Alkylene)-O(C 1-5 Alkylene)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-SH, -(C 0-3 Alkylene)-S(C 1-5 Alkyl), -(C 0-3 Alkylene)-S(C 1-5 Alkylene)-SH, -(C 0-3 Alkylene)-S(C 1-5 Alkylene)-S(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH2, -(C 0-3 Alkylene)-NH(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-OH, -(C 0-3 Alkylene)-N(C 1-5 Alkyl)-OH, -(C 0-3 Alkylene)-NH-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C 1-5 Alkyl)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-halogen, -(C 0-3 Alkylene)-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-O-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-CN, -(C 0-3 Alkylene)-NO2, -(C 0-3 Alkylene)-CHO, -(C 0-3 Alkylene)-CO-(C1-5 Alkyl), -(C 0-3 Alkylene)-COOH,-(C 0-3 Alkylene)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-NH2,-(C 0-3 Alkylene)-CO-NH(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-NH-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-N(C 1-5 Alkyl)-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-NH2,-(C 0-3 Alkylene)-SO2-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-(C 1-5 Alkyl), -(C 0-3(Alkylene)-SO-(C 1-5 alkyl), -(C 0-3 alkylene)-carbocyclic, and -(C 0-3 alkylene)-heterocyclic, where the -(C 0-3 alkylene)-carbocyclic carbocyclic moiety and the -(C 0-3 alkylene)-heterocyclic heterocyclic moiety are each independently selected from C 1-4 alkyl, halogen, -CN, -NO2, -OH, -O-(C 1-4 alkyl), -SH, -S-(C 1-4 alkyl), -NH2, -NH(C 1-4 alkyl), -N(C 1-4 alkyl)(C 1-4 alkyl), -COOH, -COO(C 1-4 alkyl), -CONH2, -CONH(C 1-4 alkyl), -CON(C 1-4 alkyl)(C 1-4 alkyl), -NHCO(C 1-4 alkyl) and -N(C 1-4 alkyl)-CO(C 1-4 alkyl) and may be substituted with one or more groups independently selected therefrom; And in the above formula, n is 0, 1, 2 or 3.

[0019] In a second aspect, the present invention relates to a polymer contained in the biosensor of the first aspect described above. In a third aspect, the present invention relates to a biosensor for measuring the glucose concentration according to the first aspect of the present invention for use in an in vivo diagnostic method.

[0020] In a fourth aspect, the present invention relates to a biosensor for measuring the glucose concentration according to the first aspect of the present invention for use in an in vivo glucose monitoring method.

[0021] In a fifth aspect, the present invention relates to a biosensor for measuring glucose concentration as described in the first aspect of the present invention, for use in in vivo diagnostic methods for hyperglycemia or hypoglycemia.

[0022] In a sixth embodiment, the present invention relates to the use of a biosensor according to the first embodiment of the present invention in an in vitro diagnostic method. In a seventh embodiment, the present invention relates to the use of a biosensor according to the first embodiment of the present invention for measuring the glucose concentration in a sample.

[0023] In an eighth aspect, the present invention relates to the use of a biosensor according to the first aspect of the present invention in an in vitro diagnostic method for hyperglycemia or hypoglycemia. In a ninth aspect, the present invention relates to the use of a polymer described in a second aspect of the present invention for producing a reagent or biosensor for monitoring blood glucose levels in a subject.

[0024] In the tenth aspect, the present invention relates to a glucose concentration-sensitive release formulation comprising the polymer described in the second aspect. In an eleventh embodiment, the present invention relates to a biosensor or glucose-sensitive release formulation for use in the treatment of glucose-dependent diseases (e.g., diabetes).

[0025] The present invention will be further illustrated with the accompanying drawings, but these should not be considered limitations. [Brief explanation of the drawing]

[0026] [Figure 1] Figure 1 shows the response of the ADAM-3%-TMAPAA sensor to changes in length at various glucose concentrations (shown). PBS pH 7.4, 37°C. [Figure 2] Figure 2 shows a comparison of pH interference between the 3APB sensor (panel A) and the ADAM sensor (panel B). [Figure 3-1]Figure 3A shows the ADAM sensor's response to glucose as TMAPAA mol% increases, and Figure 3B shows the ADAM sensor's interference with 1 mM fructose at 2.2 mM glucose as TMAPAA mol% increases. ±0.85 mM indicates the acceptable limit of interference. PBS pH 7.4, 37°C. [Figure 3-2] Figure 3C shows the interference of the ADAM sensor with 5 mM mannitol at 2.2 mM glucose as the mol% of TMAPAA increases, and Figure 3D shows the interference with 38 mM NaCl. ±0.85 mM indicates the acceptable limit of interference. PBS pH 7.4, 37°C. [Figure 3-3] Figure 3E shows the interference of the ADAM sensor with 10 mM lactate at 2.2 mM glucose as the mol% increase in TMAPAA, and Figure 3F shows the interference with 2 mM citrate. ±0.85 mM indicates the acceptable limit of interference. PBS pH 7.4 37°C. [Figure 3-4] Figure 3G shows the interference of the ADAM sensor with pH 6.9 at 2.2 mM glucose as the mol% increase in TMAPAA, and Figure 3H shows the interference with pH 7.6. ±0.85 mM indicates the acceptable limit of interference. PBS pH 7.4 37°C. [Figure 4] Figure 4 shows the response curves of the ADAM-3%-TMAPAA sensor (1-5) to various interfering substances at glucose (in mM) and 2.2 mM glucose: FRU (1 mM fructose), MAN (5 mM mannitol), LAC (10 mM lactate), CIT (2 mM citrate), NaCl (38 mM), pH 6.9, and pH 7.6. The concentrations of the tested interfering substances are sufficiently higher than those expected in vivo. [Figure 5] Figure 5 shows the effect of copolymerization of 2APB and ADAM on the interference with 5 mM mannitol (in all cases). [Figure 6]Figure 6A shows a model of a Fabry-Perot interferometer. A hydrogel constitutes the Fabry-Perot cavity. The refractive indices neff, ng, and nf are the effective refractive index of the fiber, the refractive index of the hydrogel, and the refractive index of the fluid, respectively. The distance between the two boundaries is denoted by Lg and is also called the length of the hydrogel. Figure 6B shows a typical interference pattern from a low-finesse Fabry-Perot cavity. The wavelength of light has been converted to frequency on the X-axis. The observed phase shift of 0.35 is equal to a change of 100 nm in the length of the hydrogel cavity. [Figure 7A] Figure 7A shows the change in sensor length as glucose concentration increases for the ADAM-HEAA sensor (dashed line) and the ADAM-acrylamide sensor (solid line) (Panel A shows the hydrogel containing TMAPAA). [Figure 7B] Figure 7B shows the change in sensor length as glucose concentration increases for ADAM-HEAA sensors (dashed line) and ADAM-acrylamide sensors (solid line) (Panel B shows hydrogels without TMAPAA). [Figure 8] Figure 8 shows the effect of interference from the presence of 2 mM citrate when detecting a 6 mM glucose concentration. [Figure 9A] Figure 9A shows the glucose response (in mM) of the post-modified ADAM sensor. PBS, pH 7.4, 37°C. [Figure 9B] Figure 9B shows the glucose response (expressed in mM) of a direct polymerization ADAM sensor containing 28 mol% ADAM. pH 7.4, 37°C, PBS. [Modes for carrying out the invention]

[0027] As described above, the present invention relates to a biosensor for measuring glucose concentration, wherein the biosensor is based on formula (I):

[0028] [ka]

[0029] The polymer (preferably a polymeric hydrogel) contains the portion of formula (I), and the portion of formula (I) is fixed to the polymer. In equation (I), each R is independently C 1-5 Alkyl, C 2-5 Alkenyl or C 2-5 It is an alkynyl. Preferably, each is independently C 1-5 It is alkyl. More preferably, each R is C 1-2 It is alkyl. More preferably, each R is methyl.

[0030] In equation (I), each R S Independently, C 1-5 Alkyl, C 2-5 Alkenil, C 2-5 Alkinyl, -(C 0-3 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-NH2,-(C 0-3 Alkylene)-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-OH,-(C 0-3 Alkylene)-N(C) 1-5 alkyl)-OH, -(C 0-3 Alkylene)-NH-O(C) 1-5Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-halogen, -(C 0-3 Alkylene)-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-O-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-CN,-(C 0-3 Alkylene)-NO2,-(C 0-3 Alkilen)-CHO,-(C 0-3 Alkylene)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-COOH,-(C 0-3 Alkylene)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-NH2,-(C 0-3 Alkylene)-CO-NH(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-NH-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-N(C 1-5 Alkyl)-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-NH2,-(C0-3 Alkylene)-SO2-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-(C 1-5 Alkyl), -(C 0-3 Alkilen)-SO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-Carbocyclyl, and -(C 0-3 Selected from alkylene)-heterocyclines, where the -(C) 0-3 The carbocyclyl portion of alkylene)-carbocyclyl and the -(C 0-3 The heterocycline portion of alkylene-heterocyclyl is C 1-4 Alkyl, halogen, -CN, -NO2, -OH, -O-(C 1-4 Alkyl), -SH, -S-(C 1-4 Alkyl), -NH2, -NH(C 1-4 Alkyl), -N(C 1-4 Alkyl)(C 1-4 Alkyl), -COOH, -COO(C 1-4 Alkyl), -CONH2, -CONH(C 1-4 Alkyl), -CON(C 1-4 Alkyl)(C 1-4 Alkyl), -NHCO(C 1-4 Alkyl) and -N(C 1-4 Alkyl)-CO(C 1-4 It may be substituted with one or more groups independently selected from alkyl groups.

[0031] Preferably, each R S Independently, C 1-5 Alkyl, C 2-5 Alkenil, C 2-5 Alkinyl, -(C0-3 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-NH2,-(C 0-3 Alkylene)-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-OH,-(C 0-3 Alkylene)-N(C) 1-5 alkyl)-OH, -(C 0-3 Alkylene)-NH-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-halogen, -(C 0-3 Alkylene)-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-O-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-CN,-(C 0-3 Alkylene)-NO2,-(C 0-3 Alkilen)-CHO,-(C 0-3 Alkylene)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-COOH,-(C 0-3Alkylene)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-NH2,-(C 0-3 Alkylene)-CO-NH(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-NH-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-N(C 1-5 Alkyl)-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-NH2,-(C 0-3 Alkylene)-SO2-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-SO2-(C 1-5 Alkyl), -(C 0-3 Alkylene)-SO2-(C 1-5 Alkyl), and -(C 0-3 Alkilen)-SO-(C 1-5 Selected from alkyl groups.

[0032] More preferably, each R S Independently, C 1-5 Alkyl, C 2-5 Alkenil, C 2-5 Alkinyl, -(C 0-3 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-NH2,-(C 0-3 Alkylene)-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-OH,-(C 0-3 Alkylene)-N(C) 1-5 alkyl)-OH, -(C 0-3 Alkylene)-NH-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-halogen, -(C 0-3 Alkylene)-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-O-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-CN,-(C 0-3 Alkylene)-NO2,-(C 0-3Alkilen)-CHO,-(C 0-3 Alkylene)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-COOH,-(C 0-3 Alkylene)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-NH2,-(C 0-3 Alkylene)-CO-NH(C 1-5 Alkyl), -(C 0-3 Alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-CO-O-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-NH-(C 1-5 Alkyl), -(C 0-3 Alkylene)-O-CO-N(C 1-5 Alkyl)-(C 1-5 Selected from alkyl groups.

[0033] More preferably, each R S Independently, C 1-5 Alkyl, C 2-5 Alkenil, C 2-5 Alkinyl, -(C 0-3 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-OH,-(C 0-3 Alkylene)-O(C) 1-5 Alkylene)-O(C)1-5 Alkyl), -(C 0-3 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-SH,-(C 0-3 Alkylene)-S(C) 1-5 Alkylene)-S(C) 1-5 Alkyl), -(C 0-3 Alkylene)-NH2,-(C 0-3 Alkylene)-NH(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)(C 1-5 Alkyl), -(C 0-3 Alkylene)-NH-OH,-(C 0-3 Alkylene)-N(C) 1-5 alkyl)-OH, -(C 0-3 Alkylene)-NH-O(C) 1-5 Alkyl), -(C 0-3 Alkylene)-N(C) 1-5 Alkyl)-O(C 1-5 Alkyl), -(C 0-3 Alkylene)-halogen, -(C 0-3 Alkylene)-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-O-(C 1-5 Haloalkyl), -(C 0-3 Alkylene)-CN, and -(C 0-3 Alkylene) - Selected from NO2

[0034] More preferably, each R S Independently, C 1-5 Alkyl, C 2-5 Alkenil, C 2-5 Alkinyl, -OH, -O(C) 1-5 Alkyl), -O(C 1-5 Alkylene)-OH, -O(C) 1-5 Alkylene)-O(C) 1-5 Alkyl), -SH, -S(C 1-5 Alkyl), -S(C 1-5Alkylene)-SH, -S(C 1-5 Alkylene)-S(C) 1-5 Alkyl), -NH2, -NH(C 1-5 Alkyl), -N(C 1-5 Alkyl)(C 1-5 Alkyl), -NH-OH, -N(C 1-5 Alkyl)-OH,-NH-O(C 1-5 Alkyl), -N(C 1-5 Alkyl)-O(C 1-5 Alkyl), -halogen, C 1-5 Haloalkyl, -O-(C 1-5 Selected from haloalkyl, -CN, and -NO2.

[0035] More preferably, each R S These are independently selected from -OH, -SH, -NH2, -NH-OH, -halogen, -CN, and -NO2. In formula (I), n is 0, 1, 2, or 3. Preferably, n is 0 or 1. More preferably, n is 0. The variable n is the substituent R bonded to each phenyl moiety. S It will be understood that the number of is shown. When n is 0, substituent R S Since it does not exist, the corresponding phenyl ring is unsubstituted (i.e., R S (It has hydrogen instead of ). Therefore, the phenyl ring in part of formula (I) is R S It is preferable that it is not substituted.

[0036] Furthermore, it will be understood that the portion of equation (I) can be bonded (or fixed) to the polymer via any ring carbon atom of each phenyl ring, which is reflected by the bond extending into the interior of the phenyl ring, and the wavy line (at one end of this bond) indicates the bond point of the portion of equation (I).

[0037] The manner in which the portion of formula (I) is bonded to the rest of the polymer is not particularly limited, and any chemically feasible bond is included in the present invention. For example, any of these portions can be bonded via amide bonds or reverse amide bonds, for example, via the groups -NH-CO- or -CO-NH-. Particularly preferred bonds are shown in formula (I-1) below. Further possible bond sites are shown in formula (Ib) below. Further examples of bond sites, including any of those described in the Examples section, are evident from the disclosure of methods for producing the polymer and the monomers used.

[0038] [ka]

[0039] The polymer of the present invention is of formula (Ia):

[0040] [ka]

[0041] [In the formula, R, R S And it is preferable that n includes the part as shown in equation (I). However, in one embodiment of the present invention, the polymer of the present invention is of formula (Ib):

[0042] [ka]

[0043] [In the formula, R, R S And n includes the part as in equation (I). Those skilled in the art will understand that the portion of formula (I) can be interconverted to a form containing a five-membered ring formed through the bond interaction between the nitrogen and boron atoms of the portion of formula (I). Both forms can exist in equilibrium depending on external conditions such as the surrounding medium. In particular, in an aqueous medium, these forms can be interconverted by undergoing a ring-closing reaction or a corresponding ring-opening reaction, as shown in the following scheme.

[0044] [ka]

[0045] The polymers provided according to the present invention are not particularly limited. The polymers are preferably hydrophilic, which is beneficial when the polymer comes into contact with the blood of the subject. Preferably, the polymers form polymer hydrogels; that is, the polymers are preferably polymer hydrogels. A hydrogel is generally understood as a biphasic material comprising a solid polymer that forms a preferably porous and permeable structure, and an interstitial fluid contained therein and in contact with the solid polymer. In a hydrogel, the interstitial fluid is preferably water or an aqueous solution (e.g., a liquid composition containing at least 90% (v / v) water). It will be understood that polymer hydrogels typically involve crosslinking between individual polymer molecules that form the hydrogel. Such crosslinking can be covalent or non-covalent (and consequently can be dynamic, i.e., easily moldable and deformable).

[0046] The term polymer preferably also includes polymers without dendrimers and crosslinking. The term "polymer" preferably refers to a polymer that includes crosslinking (covalent or non-covalent bonds) between individual polymer molecules.

[0047] Preferred examples of polymers (or polymer hydrogels) used in the biosensor of the present invention are described below. Generally, it is preferable that the polymer does not contain hyaluronic acid. Therefore, it is preferable that the polymer hydrogel contained in the biosensor of the present invention does not contain hyaluronic acid.

[0048] Preferably, the polymer of the present invention is a polyacrylamide-based polymer. The term “polyacrylamide-based polymer” refers to a polymer produced (or composed of) more than 50% (w / w) of acrylamide monomer, more preferably at least 60% (w / w) of acrylamide monomer, even more preferably at least 70% (w / w) of acrylamide monomer, even more preferably at least 80% (w / w) of acrylamide monomer, or even more preferably at least 90% (w / w) of acrylamide monomer. Accordingly, a polyacrylamide-based polymer can preferably be obtained by polymerization of a monomer composition in which more than 50% (w / w) (or, in increasing order of suitability, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of monomer is acrylamide monomer.

[0049] It will be understood that "acrylamide monomer" is a compound containing an acrylamide moiety, such as the one shown below.

[0050] [ka]

[0051] Such acrylamide monomers can also be called N-substituted acrylamide monomers. In principle, further substitution of the alkenyl portion of the above acrylamide monomer is possible, but it is preferable that the alkenyl portion remains unsubstituted. Therefore, we can refer to unsubstituted polyacrylamide-based polymers, which preferably consist of polymers produced (or composed of) polymers made from more than 50% (w / w) of acrylamide monomer with an unsubstituted alkenyl portion, more preferably at least 60% (w / w) of acrylamide monomer with an unsubstituted alkenyl portion, even more preferably at least 70% (w / w) of acrylamide monomer with an unsubstituted alkenyl portion, even more preferably at least 80% (w / w) of acrylamide monomer with an unsubstituted alkenyl portion, or even more preferably at least 90% (w / w) of acrylamide monomer with an unsubstituted alkenyl portion. Therefore, unsubstituted polyacrylamide-based polymers can preferably be obtained by polymerization of a monomer composition in which more than 50% (w / w) (or, in increasing order of suitability, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of monomers are unsubstituted acrylamide monomers.

[0052] The use of acrylamide monomers is beneficial because these monomers possess both the desired hydrophilicity, which facilitates the formation of hydrogels, and the ease of functionalization necessary for constructing the polymers of the present invention.

[0053] The polymer of the present invention may also be a polymethacrylamide-based polymer. The term “polymethacrylamide-based polymer” refers to a polymer produced (or composed of) preferably more than 50% (w / w) of methacrylamide monomer, more preferably at least 60% (w / w) of methacrylamide monomer, even more preferably at least 70% (w / w) of methacrylamide monomer, even more preferably at least 80% (w / w) of methacrylamide monomer, or even more preferably at least 90% (w / w) of methacrylamide monomer. Accordingly, a methacrylamide-based polymer can preferably be obtained by polymerization of a monomer composition in which more than 50% (w / w) (or, in increasing order of suitability, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of monomer is methacrylamide monomer.

[0054] It will be understood that a “methacrylamide monomer” is a compound containing a methacrylamide moiety, such as those shown below.

[0055] [ka]

[0056] Such methacrylamide monomers can also be called N-substituted methacrylamide monomers. The polymer of the present invention may also be a polyacrylate-based polymer. The term “polyacrylate-based polymer” refers to a polymer produced (or composed of) preferably more than 50% (w / w) of acrylate monomer, more preferably at least 60% (w / w) of acrylate monomer, even more preferably at least 70% (w / w) of acrylate monomer, even more preferably at least 80% (w / w) of acrylate monomer, or even more preferably at least 90% (w / w) of acrylate monomer. Accordingly, a polyacrylate-based polymer can preferably be obtained by polymerization of a monomer composition in which more than 50% (w / w) (or, in increasing order of suitability, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of monomer is acrylate monomer.

[0057] It will be understood that an "acrylate monomer" is a compound that contains an acrylate moiety, such as those shown below.

[0058] [ka]

[0059] The polymer of the present invention may also be a polymethacrylate-based polymer. The term “polymethacrylate-based polymer” refers to a polymer produced (or composed of) preferably more than 50% (w / w) of methacrylate monomer, more preferably at least 60% (w / w) of methacrylate monomer, even more preferably at least 70% (w / w) of methacrylate monomer, even more preferably at least 80% (w / w) of methacrylate monomer, or even more preferably at least 90% (w / w) of methacrylate monomer. Accordingly, a polymethacrylate-based polymer can preferably be obtained by polymerization of a monomer composition in which more than 50% (w / w) (or, in increasing order of suitability, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of monomer is methacrylate monomer.

[0060] It will be understood that a “methacrylate monomer” is a compound that contains a methacrylate moiety, such as those shown below.

[0061] [ka]

[0062] Polyacrylamide-based polymers in which the acrylamide is an unsubstituted acrylamide have been shown to be more suitable for use in the biosensors of the present invention compared to other polymers, and in particular compared to polymers based on substituted acrylamides. In particular, sensors based on unsubstituted polyacrylamide have been shown to perform better, at least in terms of sensitivity, than sensors based on substituted polyacrylamides, such as HEAA (N-(2-hydroxyethyl)acrylamide). Accordingly, for the same monomer ratio, the change in length of the ADAM-acrylamide sensor at 20 mM glucose has been shown to be approximately 1.6 times greater than that of the ADAM-HEAA sensor (see Example 2 for further details).

[0063] Therefore, the polymer is preferably an unsubstituted polyacrylamide-based polymer. In this specification, “measurement of glucose concentration” may include measuring the absolute value of the glucose concentration in a liquid (e.g., blood) in contact with a biosensor. However, measurement of glucose concentration also refers to determining the glucose concentration in comparison to a threshold. For example, measurement of glucose concentration may also include determining whether the glucose concentration is below or above a threshold, or whether the concentration falls within a reference range. This is particularly useful in clinical settings, for example, when blood glucose levels fall below a certain critical threshold requiring intervention by a physician.

[0064] The measurement of glucose concentration using the biosensor of the present invention is based on the fact that the volume of a polymer changes when it comes into contact with a glucose-containing liquid, in other words, when it comes into contact with glucose. The present invention is at least in part based on the surprising discovery that the polymer of the present invention decreases in volume when it comes into contact with glucose. Thus, when the polymer (or preferably a polymer hydrogel) comes into contact with a glucose-free liquid (which can also be called a liquid containing glucose below the minimum detection limit concentration or below a certain threshold), in other words, in the absence of glucose, the volume of the polymer (or polymer hydrogel) remains unchanged because the boronic acid-based glucose-binding portion of the polymer (or polymer hydrogel) does not form any new interactions. Then, when exposed to a glucose-containing liquid (i.e., containing glucose at a detectable concentration or a concentration of glucose above a certain threshold), in other words, in the presence of glucose, a change in the volume of the polymer occurs as the boronic acid-based glucose-binding portion binds to glucose. The change in the volume of the polymer (or polymer hydrogel) that can be observed when transitioning from a glucose-free liquid to a glucose-containing liquid, in other words, when transitioning from the absence of glucose to the presence of glucose, is a decrease in the volume of the polymer (or polymer hydrogel), i.e., shrinkage. Although I don't want to be constrained by theory, the shrinkage of the polymer upon contact with glucose is thought to be due to the formation of a stoichiometric 2:1 complex between the part of equation (I) and glucose, as shown in the following scheme.

[0065] [ka]

[0066] Thus, preferably, in the biosensor for measuring glucose concentration of the present invention, the polymer shrinks when it comes into contact with glucose. The degree of contraction is preferably substantially linearly proportional to the glucose concentration. Here, the degree of contraction is preferably understood as |ΔL / L|, i.e., the absolute value obtained from ΔL / L. In the formula, ΔL is the change in the linear dimension of the polymer (or polymer hydrogel), and L is the total linear dimension of the polymer (or polymer hydrogel). Accordingly, biosensors containing the polymers described herein preferably exhibit a substantially linear relationship, more preferably a linear relationship, between glucose concentration and ΔL / L. In this specification, the term “substantially linear” preferably means within the expected value of the linear relationship, particularly within a tolerance of 10% (i.e., within ±10% of the linear relationship), more preferably within a tolerance of 5%, and even more preferably within a tolerance of 2%.

[0067] As will be apparent to those skilled in the art, if the degree of contraction depends on the glucose concentration, it becomes possible to detect changes in glucose concentration. For example, by creating a baseline curve of the degree of expansion / contraction (ΔL / L) measured at several known glucose concentrations and comparing the measured degree of expansion / contraction with the baseline curve, the glucose concentration can be determined.

[0068] Whenever the term “about” is used in relation to a numerical value in this specification, it preferably refers to ±10% of the indicated numerical value, more preferably ±5%, even more preferably ±2%, even more preferably ±1% of the indicated numerical value, and most preferably the exact indicated numerical value. When the term “about” is used in relation to the endpoint of a range, it preferably refers to the range from -10% of the lower endpoint of the indicated numerical value to +10% of the upper endpoint of the indicated numerical value, more preferably the range from -5% of the lower endpoint to +5% of the upper endpoint, even more preferably the range from -2% of the lower endpoint to +2% of the upper endpoint, even more preferably the range from -1% of the lower endpoint to +1% of the upper endpoint, and most preferably the range defined by the exact numerical values ​​of the lower and upper endpoints. Note that the numerical value may also be a ratio, for example, the ratio used to represent the composition of the polymer of the present invention as described above. A ratio can be written as a division operation of two numbers, but it can also be expressed as a single number obtained from such division. For example, a ratio of 1:2 can also be expressed as a ratio of 0.5. The aforementioned ±x% deviation can be applied to a single corresponding number (obtained from the division operation), and the endpoints of the resulting range can be converted back to their corresponding ratios. For example, a ratio of 1:2 ± 10% can also be expressed as a ratio of 0.5 ± 10%, or as a ratio of 0.45 to 0.55 (corresponding to a ratio of 1:1.82 to 1:2.22).

[0069] Furthermore, the determination / detection of polymer volume changes can also be performed by measuring another relevant value, particularly a proxy value (or surrogate value) that depends on the volume of the polymer, such as osmotic pressure (or change in osmotic pressure). Accordingly, the present invention also relates to a biosensor for glucose concentration measurement (as described herein), in which the measurement of glucose concentration is based on glucose concentration-sensitive changes in osmotic pressure within a polymer (or polymer hydrogel as preferred in the present invention) contained in the biosensor.

[0070] The volume change, i.e., expansion / contraction, of a polymer (polymeric hydrogel) in response to contact with glucose is reversible and can be measured (and thus monitored in real time) by various signaling mechanisms, including electrochemical, mechanical, and optical techniques (e.g., Fabry-Perot interferometry or refractive index measurement). The refractive index of the polymer may change simply as a result of the volume change (i.e., as the polymer expands, it becomes more dilute and therefore its refractive index decreases) and / or because the sample molecule (analyte) binds to the polymer chain.

[0071] Polymers (preferably polymeric hydrogels) have been described above. According to established practice in the art, polymers can be defined in terms of the monomers polymerized, in particular, according to the method of production. Therefore, it is preferable that the polymer (or polymeric hydrogel) be obtained by a polymerization reaction of a composition comprising the following:

[0072] (i) An acrylamide monomer containing a boronic acid-based glucose-binding moiety of formula (I). It will be understood that the boronic acid-based glucose-binding moiety in acrylamide monomer (i) is bonded to the rest of the monomer through a bond site, as shown above. Note that some formulas showing preferred embodiments of the boronic acid-based glucose-binding moiety include an -NH-CO- moiety. In the corresponding acrylamide monomers (i) or (ii), this -NH-CO- moiety preferably forms part of the acrylamide moiety. That is, the -CO- of the -NH-CO- moiety preferably bonds to an ethylene group (-CH=CH2) to form the acrylamide moiety -NH-CO-CH=CH2.

[0073] (i) is,

[0074] [ka]

[0075] [In the formula, R, R S It is particularly preferable that n is as shown in equation (I). However, in one form, (i) is,

[0076] [ka]

[0077] [In the formula, R, R S And n are as shown in formula (I). Such compounds are known from the literature (Chem.Commun., 2011, 47, 8169-8171, DOI:10.1039 / c1cc11096a).

[0078] Preferably, the composition used in the polymerization reaction further comprises the following: (ii) Acrylamide monomers that do not contain the boronic acid portion, and (iii) Acrylamide monomer as a crosslinking agent.

[0079] Acrylamide monomers that do not contain the boronic acid portion are:

[0080] [ka]

[0081] A monomer containing, preferably containing exactly one copy of such a moiety, but not containing any boronic acid moiety. Such monomers are not particularly limited, but are preferably unsubstituted acrylamide monomers (where the above moiety is bonded to hydrogen) or substituted acrylamide monomers where the above moiety is bonded to, for example, a hydroxyalkyl group or an ethylene glycol oligomer. Exemplary preferred monomers (ii) are selected from the following monomers.

[0082] [ka]

[0083] In the formula, q is an integer from 0 to 10. Preferably, q is an integer from 2 to 5, and more preferably, q is 2 or 5. One type of monomer (ii) or two or more types (for example, two or three types) of monomer (ii) may be present in the composition, and the polymer of the present invention can be obtained by its polymerization.

[0084] The acrylamide monomer (iii) of the crosslinking agent preferably has the following moiety:

[0085] [Chemical formula]

[0086] contains at least 2 copies, preferably exactly 2 copies of the above moiety. Further preferably, it does not contain a boronic acid moiety. Preferably, the acrylamide monomer (iii) of the crosslinking agent

[0087] [Chemical formula]

[0088] [In the formula, q is an integer from 0 to 10, preferably q is an integer from 2 to 5, and more preferably q is 2 or 5] and contains (or is) a monomer selected therefrom. Preferably, in the case of the polymer (more preferably, the polymer hydrogel) of the present invention, the acrylamide monomer (i) constitutes 6 to 30 mol% of the acrylamide-based component in the composition used for the polymerization reaction. More preferably, the acrylamide monomer (i) constitutes 10 to 14 mol% of the acrylamide-based component in the composition used for the polymerization reaction. However, the amount of the acrylamide monomer (I) may be more, and constitutes 15 to 24 mol%, preferably 16 to 20 mol% of the acrylamide-based component in the composition used for the polymerization reaction.

[0089] Preferably, in the polymer of the present invention, monomer (ii) constitutes 60 to 94 mol% of the acrylamide-based component in the composition used in the polymerization reaction, preferably 66 to 90 mol%, and more preferably 74 to 84 mol%.

[0090] Preferably, in the polymer of the present invention, the crosslinking agent acrylamide monomer (iii) constitutes 0.1 to 4 mol% of the acrylamide base component in the composition used in the polymerization reaction. More preferably, in the polymer of the present invention, the crosslinking agent acrylamide monomer (iii) constitutes 1 to 4 mol% of the acrylamide base component in the composition used in the polymerization reaction.

[0091] In an alternative embodiment of the present invention, the polymer (or polymeric hydrogel) can be obtained by a polymerization reaction of a composition comprising the following: (ia) A nonacrylamide monomer containing a boronic acid-based glucose-binding moiety of formula (I). Here, a nonacrylamide monomer is understood to be a monomer that does not contain the -NH-CO-CH=CH2 moiety.

[0092] Accordingly, in this alternative embodiment of the present invention, the polymer (or polymer hydrogel) can be obtained by the above polymerization reaction in which (i) is replaced with (ia). In this alternative embodiment of the present invention, (ia) is

[0093] [ka]

[0094] [In the formula, R, R S It is preferable that n and are as shown in formula (I). Such monomers are known from the literature (J.Am.Chem.Soc.2009,131,13908-13909,doi:10.1021 / ja905652w).

[0095] In a further alternative embodiment, (ia) is an acrylate monomer. The polymers of the present invention, and, if applicable, polymeric hydrogels, can be produced according to the methods described in the above section and the section on examples.

[0096] Therefore, it is preferable to incorporate the portion of formula (I) into a monomer (one or more) and then copolymerize them to form a polymer of the present invention (preferably a polymeric hydrogel) containing the portion. However, the present invention also includes embodiments in which the portion of formula (I) is incorporated into a pre-fabricated polymer through modification of the polymer. One exemplary method of such modification is a Michael-like addition reaction of an -SH group present as a thiolactone in the pre-formed polymer to the acrylamide moiety.

[0097] The polymers of the present invention, in particular the polymer hydrogels of the present invention, exhibit beneficial properties as discussed above and as shown in the examples, and are therefore particularly suitable for use in the biosensors of the present invention for measuring (or sensing) glucose concentration.

[0098] It is particularly preferable that the degree of shrinkage of the polymer (or high molecular hydrogel) in the presence of glucose is substantially independent of pH values ​​in the range of approximately pH 6.9 to approximately pH 7.6. This pH range is particularly relevant in applications in the intensive care unit (ICU), where the treated patient may exhibit not only a normal blood pH range, i.e., approximately 7.4 to approximately 7.6, but also a wider range of blood pH values, including pathological pH values.

[0099] In this specification, the expression “substantially independent of pH value” within a particular pH range preferably means that within the said pH range, the maximum change / variation in the degree of expansion / contraction is 15% (understood as ΔL / L = 15%) (depending on sensitivity (0-20 mM glucose)), more preferably 10%, and even more preferably 5%. The term “substantially independent” also includes specific references to “independent” in a narrower sense.

[0100] The biosensors and polymers of the present invention may include further parts and further monomers that may provide additional benefits. These are described below in the specific embodiments of the biosensors of the present invention disclosed.

[0101] In a first specific embodiment of the biosensor of the present invention, the polymer further includes a portion that is positively charged at pH=7.4. In this specification, a positively charged portion is understood to be a portion whose net charge at pH=7.4 is greater than zero. Therefore, a zwitterionic portion containing both positive and negative charges is not considered positively charged because its net charge is neutral.

[0102] Preferably, in this first particular embodiment, the polymer further comprises a portion containing an amino (or ammonium) group that is positively charged at pH=7.4. An example of such a group is the primary amino group -NH2 (in its ammonium form, the structure -NH3). + (having), secondary amino group -NH(C 1-5 Alkyl), for example, -NHCH3 (in its ammonium form, the structure is -(NH2(C 1-5 Alkyl)) + For example, -(NH2CH3) + (having), or tertiary amino group-N(C 1-5 Alkyl)2, for example -N(CH3)2(in its ammonium form, the structure is -(NH(C 1-5 Alkyl)2) + (and so on.)

[0103] Examples of quaternary ammonium groups are -N(C 1-5 alkyl)3 + and, for example, -N(CH3)3 + is possible. Thus, preferably, in this first specific embodiment, the polymer further comprises a moiety containing -NH2, -NH3 + , -NH(C 1-5 alkyl), -(NH2(C 1-5 alkyl)) + , -N(C 1-5 alkyl)2, -(NH(C 1-5 alkyl)2) + , and -N(C 1-5 alkyl)3 + selected from the groups.

[0104] The positively charged moiety is preferably located within the monomer used in the polymerization reaction (or, in other words, the positively charged moiety is preferably obtained / can be obtained by using a corresponding monomer as described below, for example, in the polymerization reaction). Preferably, in this specific embodiment, the monomer is selected from the following.

[0105]

Chemical formula

[0106] However, the present invention also includes an embodiment in which the above-mentioned positively charged moiety is introduced by a reaction occurring after the polymerization reaction. In other words, the positively charged moiety can also be introduced into the polymer by modifying the polymer.

[0107] Preferably, in this first specific embodiment of the biosensor of the present invention, the polymer further comprises a moiety containing a quaternary ammonium group. The moiety containing a quaternary ammonium group has the formula (II):

[0108]

Chemical formula

[0109] It could be that part. The portion containing the quaternary ammonium group, particularly the portion of formula (II), is bonded to the polymer of the present invention in the same manner as described above for the portion of formula (I). Therefore, in this first particular embodiment, the portion containing the quaternary ammonium group is preferably of formula (IIa):

[0110] [ka]

[0111] This is the relevant part. Biosensors of the present invention that include a moiety containing a quaternary ammonium group have been shown to have properties beneficial for glucose sensing. Accordingly, a first particular aspect of the present invention is at least in part based on the discovery that the selectivity of the biosensor of the present invention for glucose is increased when the polymer of the present invention further includes a quaternary ammonium group, for example, a moiety of formula (II), compared to common interfering substances (such as, but not limited to, fructose, mannitol, lactate, and citrate). Surprisingly, in the present invention, a ratio of monomer containing a quaternary ammonium group to monomer containing a boronic acid moiety of 1:2 to 1:7, preferably 1:3 to 1:6, and more preferably 1:4 to 1:5 is found to be particularly beneficial because it is sufficient to substantially suppress interference from fructose, mannitol, lactate, and citrate. A further beneficial ratio of monomer containing a quaternary ammonium group to monomer containing a boronic acid moiety is 1:5 to 1:7. This is further illustrated in Figure 3.

[0112] Therefore, it is particularly beneficial, preferably, that the ratio of monomers containing a positively charged portion at pH=7.4 (including any specific and preferred such portions disclosed herein) to monomers containing a boronic acid portion is 1:2 to 1:7, preferably 1:3 to 1:6, and more preferably 1:4 to 1:5, as this is sufficient to substantially suppress interference from fructose, mannitol, lactate, and citrate. A further beneficial ratio of monomers containing a positively charged portion at pH=7.4 (including any specific and preferred such portions disclosed herein) to monomers containing a boronic acid portion is 1:5 to 1:7.

[0113] In this specification, when interference from a particular substance is substantially suppressed, the readout is substantially independent of the concentration of such substance. In this specification, the expression “substantially independent of the concentration” of a particular substance within a particular concentration range preferably means that within the said concentration range, the maximum change / variation in the degree of expansion / contraction is observed to be 15% (understood as ΔL / L = 15%) (depending on sensitivity (0-20 mM glucose)), more preferably 10%, and even more preferably 5%. The term “substantially independent” also includes specific references to “independent” in a narrower sense.

[0114] Preferably, in this first particular embodiment of the present invention, the portion of formula (II) is introduced into the polymer through polymerization of a reaction mixture further comprising, in addition to the above components, a monomer containing the portion of formula (iv) (II). Preferably, (iv) is

[0115] [ka]

[0116] That is the case. Preferably, in this first specific embodiment of the present invention, for the polymer (more preferably a polymeric hydrogel) of the present invention, the acrylamide monomer (iv) constitutes 0.1 to 6 mol% of the acrylamide-based component in the composition / reaction mixture used in the polymerization reaction. More preferably, the acrylamide monomer (iv) constitutes 2 to 4 mol% of the acrylamide-based component in the composition used in the polymerization reaction.

[0117] As described herein, the interference suppression effect has been shown to depend on the presence of a positive charge in the monomer. The addition of TMAPPA provides a beneficial effect on the resulting glucose sensor by reducing the interference of fructose, mannitol, and citrate. Since this effect has been further shown to originate from a positive charge on the molecule, similar cationic monomers are expected to provide similar benefits to TMAPPA.

[0118] In a second specific embodiment of the present invention, the polymer is further of formula (III):

[0119] [ka]

[0120] This includes the part. Accordingly, this second particular aspect of the present invention is at least in part based on the remarkable discovery that the biosensor of the present invention exhibits suppressed sensitivity to mannitol when the portion of formula (II) is present in addition to the portion of formula (I). Preferably, the ratio of the portion of formula (III) to the portion of formula (I) is 1:10 to 1:5, more preferably 1:8 to 1:6.

[0121] Preferably, in this second particular embodiment, the portion of formula (III) is introduced into the polymer through polymerization of a reaction mixture that further includes a monomer comprising the portion of formula (III) in addition to the aforementioned components. Preferably, monomer (v) is of the following formula

[0122] [ka]

[0123] It is a compound of [the compound]. Preferably, in this second specific embodiment of the present invention, for the polymer of the present invention (more preferably a polymeric hydrogel), the acrylamide monomer (v) constitutes 1 to 5 mol% of the acrylamide base component in the composition used in the polymerization reaction. More preferably, the acrylamide monomer (v) constitutes 2 to 4 mol% of the acrylamide base component in the composition used in the polymerization reaction. Even more preferably, the acrylamide monomer (v) constitutes about 3 mol% of the acrylamide base component in the composition used in the polymerization reaction. In this second specific embodiment of the present invention, preferably, the acrylamide monomers (i) and (v) constitute about 25 mol% and about 3 mol%, respectively.

[0124] The definitions below apply to this specification and the entirety of the claims unless otherwise specified. The term "hydrocarbon group" refers to a group composed of carbon atoms and hydrogen atoms. The term "alicyclic" is used in relation to cyclic groups, indicating that the corresponding cyclic group is non-aromatic.

[0125] In this specification, the term “alkyl” refers to a monovalent saturated acyclic (non-cyclic) hydrocarbon group, which may be linear or branched. Therefore, the “alkyl” group does not include carbon-carbon double bonds or carbon-carbon triple bonds. 1-5 "Alkyl" refers to an alkyl group having 1 to 5 carbon atoms. Examples of preferred alkyl groups include methyl, ethyl, propyl (e.g., n-propyl or isopropyl), or butyl (e.g., n-butyl, isobutyl, sec-butyl, or tert-butyl). Unless otherwise defined, the term "alkyl" preferably refers to C 1-4 This refers to alkyl, more preferably methyl or ethyl, and even more preferably methyl.

[0126] In this specification, the term “alkenyl” refers to a monounsaturated acyclic hydrocarbon group that may be linear or branched, containing one or more (e.g., one or two) carbon-carbon double bonds but no carbon-carbon triple bonds. 2-5 "Alkenyl" refers to an alkenyl group having 2 to 5 carbon atoms. Preferred examples of alkenyl groups are ethenyl, propenyl (e.g., propa-1-en-1-yl, propa-1-en-2-yl, or propa-2-en-1-yl), butenyl, butadienyl (e.g., buta-1,3-dien-1-yl or buta-1,3-dien-2-yl), pentenyl, or pentadienyl (e.g., isoprenyl). Unless otherwise defined, the term "alkenyl" preferably means C 2-4 This refers to Alkenil.

[0127] In this specification, the term “alkynyl” refers to a monounsaturated acyclic hydrocarbon group that may be linear or branched, comprising one or more (e.g., one or two) carbon-carbon triple bonds and optionally one or more (e.g., one or two) carbon-carbon double bonds. 2-5 "Alkynyl" refers to an alkynyl group having 2 to 5 carbon atoms. Suitable examples of alkynyl groups are ethynyl, propynyl (e.g., propargyl), or butynyl. Unless otherwise defined, the term "alkynyl" preferably means C 2-4 This refers to alkinyl.

[0128] In this specification, the term "alkylene" refers to an alkanediyl group, i.e., a divalent saturated acyclic hydrocarbon group which may be linear or branched. 1-5 The term "alkylene" refers to an alkylene group having 1 to 5 carbon atoms, and the term "C 0-3 "Alkylene" is a covalent bond (corresponding to option "C0 alkylene") or C 1-3This indicates the presence of alkylene. Suitable examples of alkylene groups are methylene (-CH2-), ethylene (e.g., -CH2-CH2- or -CH(-CH3)-), propylene (e.g., -CH2-CH2-CH2-, -CH(-CH2-CH3)-, -CH2-CH(-CH3)-, or -CH(-CH3)-CH2-), or butylene (e.g., -CH2-CH2-CH2-CH2-). Unless otherwise defined, the term “alkylene” preferably refers to C 1-4 Alkylenes (especially straight-chain C) 1-4 This refers to (including alkylene), more preferably methylene or ethylene, and even more preferably methylene.

[0129] In this specification, the term “carbocyclyl” refers to a hydrocarbon ring group including monocyclic rings, as well as bridging rings, spiro rings, and / or fused ring systems (which may consist of, for example, two or three rings), wherein the ring group may be saturated, partially unsaturated (i.e., unsaturated but not aromatic), or aromatic. Unless otherwise defined, “carbocyclyl” preferably refers to an aryl, cycloalkyl, or cycloalkenyl.

[0130] In this specification, the term “heterocyclyl” refers to a ring group that includes a monocyclic ring as well as a bridging ring, a spiro ring, and / or a fused ring system (which may consist of, for example, two or three rings), wherein the ring group contains one or more (for example, one, two, three, or four) ring heteroatoms independently selected from O, S, and N, and the remaining ring atoms are carbon atoms. Here, one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may be oxidized, and one or more carbon ring atoms may also be oxidized (i.e., form an oxo group). Furthermore, the ring group may be saturated, partially unsaturated (i.e., unsaturated but not aromatic), or aromatic. For example, each heteroatom-containing ring contained in the ring group may contain one or two oxygen atoms and / or one or two sulfur atoms (which may be oxidized) and / or one, two, three or four nitrogen atoms (which may be oxidized), provided that the total number of heteroatoms in the corresponding heteroatom-containing ring is 1 to 4, and the corresponding heteroatom-containing ring contains at least one carbon ring atom (which may be oxidized). Unless otherwise defined, “heterocyclyl” preferably refers to a heteroaryl, heterocycloalkyl, or heterocycloalkenyl.

[0131] In this specification, the term “aryl” refers to an aromatic hydrocarbon ring group comprising a monocyclic aromatic ring and a bridged and / or fused ring system containing at least one aromatic ring (e.g., a ring system composed of two or three fused rings, at least one of which is aromatic; or a bridged ring system composed of two or three rings, at least one of which is aromatic). If the aryl is a bridged and / or fused ring system containing at least one non-aromatic ring (e.g., a saturated or unsaturated alicyclic ring) in addition to one or more aromatic rings, one or more carbocyclic atoms in each non-aromatic ring may be oxidized (i.e., form an oxo group). "Aryl" may refer to, for example, phenyl, naphthyl, dialinyl (i.e., 1,2-dihydronaphthyl), tetralinyl (i.e., 1,2,3,4-tetrahydronaphthyl), indanyl, indenyl (e.g., 1H-indenyl), anthracenyl, phenantrenyl, 9H-fluorenyl, or azlenyl. Unless otherwise defined, "aryl" preferably has 6 to 14 ring atoms, more preferably 6 to 10 ring atoms, and even more preferably phenyl or naphthyl, most preferably phenyl.

[0132] In this specification, the term “heteroaryl” means an aromatic ring group comprising a bridging ring and / or fused ring system (e.g., a ring system composed of two or three fused rings, at least one of which is aromatic; or a bridging ring system composed of two or three rings, at least one of which is aromatic) that includes a monocyclic aromatic ring and at least one other aromatic ring, wherein the aromatic ring group comprises one or more (e.g., one, two, three, or four) ring heteroatoms independently selected from O, S, and N, and the remaining ring atoms are carbon atoms. Here, one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may be oxidized, and one or more carbocyclic atoms may also be oxidized (i.e., to form an oxo group). For example, each heteroatom-containing ring contained in the aromatic ring group may contain one or two oxygen atoms and / or one or two sulfur atoms (which may be oxidized) and / or one, two, three or four nitrogen atoms (which may be oxidized), provided that the total number of heteroatoms in the corresponding heteroatom-containing ring is 1 to 4, and at least one carbon ring atom (which may be oxidized) is present in the corresponding heteroatom-containing ring. "Heteroaryls" include, for example, thienyl (i.e., thiophenyl), benzo[b]thienyl, naphtho[2,3-b]thienyl, thianthrenyl, furyl (i.e., furanyl), benzofuranyl, isobenzofuranyl, chromanyl, clomenyl (e.g., 2H-1-benzopyranyl or 4H-1-benzopyranyl), isoclomenyl (e.g., 1H-2-benzopyranyl), chromonyl, xanthenyl, phenoxathinyl, pyrrolyl (e.g., 1H-pyrrolyl), imidazolyl, pyrazolyl, pyridyl (i.e., pyridinyl); For example, 2-pyridyl, 3-pyridyl, or 4-pyridyl), pyrazinyl, pyrimidinyl, pyridadinyl, indolyl (e.g., 3H-indolyl), isoindolyl, indazolyl, indolidinyl, prinyl, quinolyl, isoquinolyl, phthalazinyl, naphthylidinyl, quinoxalinyl, cinnolinyl, pteridinyl, carbazolyl, β-carbolinyl, phenanthrolinyl, acridinyl, perimidinyl, phenanthrolinyl (e.g., [1,10]phenanthrolinyl, [1,7]phenanthrolinyl, or [4,7) Phenanthrolinyl), phenazinyl, thiazolyl, isothiazolyl, phenothiazinyl, oxazolyl, isoxazolyl, oxadiazolyl (e.g., 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl (i.e., flazanil), or 1,3,4-oxadiazolyl), thiadiazolyl (e.g., 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, or 1,3,4-thiadiazolyl) , phenoxadinyl, pyrazolo[1,5-a]pyrimidinyl (e.g., pyrazolo[1,5-a]pyrimidine-3-yl), 1,2-benzoisoxazole-3-yl, benzothiazolyl, benzothiadiazolyl, benzoxazolyl, benzisoxazolyl, benzimidazolyl, benzo[b]thiophenyl (i.e., benzothienyl), triazolyl (e.g., 1H-1,2,3-triazolyl, 2H-1,2, 3-Triazolyl, 1H-1,2,4-Triazolyl, or 4H-1,2,4-Triazolyl), benzotriazolyl, 1H-Tetrazolyl, 2H-Tetrazolyl, triazinyl (e.g., 1,2,3-Triadinyl, 1,2,4-Triadinyl, or 1,3,5-Triadinyl), flo[2,3-c]pyridinyl, dihydroflopyridinyl (e.g., 2,3-dihydrofloo[2,3-c]pyridinyl or 1,3-di Hydroflo[3,4-c]pyridinyl), imidazopyridinyl (e.g., imidazo[1,2-a]pyridinyl or imidazo[3,2-a]pyridinyl), quinazolinyl, thienopyridinyl, tetrahydrothienopyridinyl (e.g., 4,5,6,7-tetrahydrothieno[3,2-c]pyridinyl), dibenzofuranyl, 1,3-benzodioxolyl, benzodioxanyl (e.g., 1,3-benzodioxanyl or 1,This can refer to 4-benzodioxanyl, or coumarinyl. Unless otherwise defined, “heteroaryl” is preferably a monocyclic or fused ring system containing one or more (e.g., one, two, three, or four) ring heteroatoms independently selected from O, S, and N, with one or more S ring atoms (if present) and / or one or more N ring atoms (if present) being oxidized, and one or more carbocyclic atoms also being oxidized. More preferably, “heteroaryl” is a monocyclic ring containing one or more (e.g., one, two, or three) ring heteroatoms independently selected from O, S, and N, with one or more S ring atoms (if present) and / or one or more N ring atoms (if present) being oxidized, and one or more carbocyclic atoms also being oxidized.

[0133] In this specification, the term “cycloalkyl” refers to a saturated hydrocarbon ring group and includes monocyclic rings as well as bridging rings, spiro rings and / or fused ring systems (e.g., which may consist of two or three rings; e.g., fused ring systems consisting of two or three fused rings). “Cycloalkyl” can refer to, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, dekalinyl (i.e., decahydronaphthyl), or adamantyl. Unless otherwise defined, “cycloalkyl” preferably refers to C 3-11 This refers to a cycloalkyl group, and more preferably C 3-7 This refers to cycloalkyl groups. Particularly preferred "cycloalkyl groups" are monocyclic saturated hydrocarbon rings having 3 to 7 ring members (e.g., cyclopropyl or cyclohexyl).

[0134] In this specification, the term “heterocycloalkyl” refers to a saturated ring group that includes a monocyclic ring as well as a bridging ring, a spiro ring, and / or a fused ring system (for example, which may consist of two or three rings; for example, a fused ring system consisting of two or three rings), wherein the ring group contains one or more (for example, one, two, three, or four) ring heteroatoms independently selected from O, S, and N, and the remaining ring atoms are carbon atoms. Here, one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may be oxidized, and furthermore, one or more carbocyclic atoms may also be oxidized (i.e., forming an oxo group). For example, each heteroatom-containing ring contained in the saturated ring group may contain one or two oxygen atoms and / or one or two sulfur atoms (which may be oxidized) and / or one, two, three or four nitrogen atoms (which may be oxidized), provided that the total number of heteroatoms in the corresponding heteroatom-containing ring is 1 to 4, and the corresponding heteroatom-containing ring contains at least one carbocyclic atom (which may be oxidized). Examples of "heterocycloalkyl" include azilidinyl, azetidinyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, azepanyl, diazepanyl (e.g., 1,4-diazepanyl), oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolidinyl, morpholinyl (e.g., morpholine-4-yl), thiomorpholinyl (e.g., thiomorpholine-4-yl), oxazepanyl, oxylan This can refer to 1,4-dioxolanil, 1,3-tetrahydrofuranil, 1,3-dioxolanil, tetrahydropyranil, 1,4-dioxanil, oxepanil, thiranil, thietanil, tetrahydrothiophenyl (i.e., thioranil), 1,3-dithioranil, thianil, 1,1-dioxothianil, thiepanil, decahydroquinolinil, decahydroisoquinolinil, or 2-oxa-5-azabicyclo[2.2.1]hepta-5-yl.Unless otherwise defined, “heterocycloalkyl” preferably refers to a saturated ring group which is a 3- to 11-membered monocyclic ring or fused ring system (e.g., a fused ring system composed of two fused rings), wherein the ring group contains one or more (e.g., one, two, three, or four) ring heteroatoms independently selected from O, S, and N, and one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may be oxidized, and one or more carbocyclic atoms may also be oxidized. More preferably, “heterocycloalkyl” refers to a saturated monocyclic ring group which contains one or more (e.g., one, two, or three) ring heteroatoms independently selected from O, S, and N, and one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may be oxidized, and one or more carbocyclic atoms may also be oxidized.

[0135] In this specification, the term “halogen” refers to fluoro(-F), chloro(-Cl), bromo(-Br), or iodine(-I). The terms “halogen” and “halo” are interchangeable.

[0136] In this specification, the term “haloalkyl” refers to an alkyl group substituted with one or more (preferably 1 to 6, more preferably 1 to 3) halogen atoms, wherein the halogen atoms are independently selected from fluoro, chloro, bromo, and iodine, and preferably all are fluoro atoms. It will be understood that the maximum number of halogen atoms is limited by the number of available bonding sites and therefore depends on the number of carbon atoms in the alkyl portion of the heteroalkyl group. “Haloalkyl” can refer to, for example, -CF3, -CHF2, -CH2F, -CF2-CH3, -CH2-CF3, -CH2-CHF2, -CH2-CF2-CH3, -CH2-CF2-CF3, or -CH(CF3)2. A particularly preferred “haloalkyl” group is -CF3.

[0137] The terms “joint” and “covalent bond” are used synonymously in this specification unless explicitly indicated otherwise or unless inconsistent with the context. In this specification, the terms “optional,” “optionally,” and “may” indicate that the indicated feature may or may not be present. Whenever “optional,” “optionally,” and “may” are used, the present invention specifically relates to both possibilities, i.e., the presence or absence of the corresponding feature. For example, the expression “X is optionally substituted with Y” (or “X may be substituted with Y”) means that X is either substituted with Y or not. Similarly, where a component of a composition is indicated as “optional,” the present invention specifically relates to both possibilities, i.e., the presence (contained in the composition) or the absence of the corresponding component in the composition.

[0138] In this specification, various groups are referred to as "optionally substituted (or may be substituted)." Generally, these groups may have one or more substituents, e.g., one, two, three, or four substituents. It will be understood that the maximum number of substituents is limited by the number of binding sites available in the substituted portion. Unless otherwise defined, the "optionally substituted (or may be substituted)" groups referred to herein preferably have no more than two substituents, in particular just one substituent. Furthermore, unless otherwise defined, it is preferable that there are no substituents, i.e., the corresponding group is unsubstituted.

[0139] Those skilled in the art will understand that the substituents contained in the compounds of the present invention may be bonded to the remainder of each compound via several different positions of the corresponding specific substituent. Unless otherwise defined, preferred bonding positions of various specific substituents are as shown in the examples.

[0140] In this specification, unless otherwise explicitly indicated or unless inconsistent with the context, the terms “a,” “an,” and “the” are used interchangeably with “one or more” and “at least one.”

[0141] Wherever a numerical range is provided / disclosed herein, it should be understood that all values ​​and subranges encompassed by that numerical range are also included within the scope of the present invention. Accordingly, the present invention relates specifically and individually to each value that falls within the numerical range disclosed herein, in particular to each individual integer value that falls within the corresponding numerical range, and to each subrange encompassed within the numerical range disclosed herein.

[0142] In this specification, the term “comprising” (or “comprise,” “comprises,” “contain,” “contains,” or “containing”) means “particularly containing,” that is, “particularly containing among any further elements…” unless otherwise explicitly indicated or inconsistent with the context. In addition, the term also includes the narrower meanings of “essentially consisting of” and “consisting of.” For example, the term “A contains B and C” means “A contains B and C in particular,” and A may contain any further elements (including, for example, “A contains B, C and D”), but the term also includes the meanings of “A essentially consists of B and C” and “A consists of B and C” (i.e., A does not contain any other components besides B and C).

[0143] In this specification, the terms “subject” or “patient” refer to an animal, preferably a mammal (e.g., human or non-human mammal). Most preferably, “subject” or “patient” is human (e.g., human male or human female).

[0144] The measurement of glucose concentration using the biosensor of the present invention typically requires the measurement of volume changes detected through the measurement of a polymer (or polymer hydrogel), particularly through the measurement of changes in the length of the polymer / polymer hydrogel. The corresponding measurement can be carried out using the method described in US 7,602,498 B2, the said patent is incorporated herein by reference in its entirety. In particular, to measure the length of the polymer hydrogel, the change in the length of the hydrogel induced by various glucose concentrations can be monitored using the reflection interference spectrum from the hydrogel. The hydrogel is placed at the tip of a cut single-mode fiber. The relative length change of the hydrogel is measured from the initial absolute length L g (t=t1) The measurement is continuously monitored. The length of the hydrogel is measured from the center of the cross-section to the tip of the hydrogel along a straight path perpendicular to the cross-section.

[0145] The reflection spectrum of the hydrogel is approximated by a sine wave with DC components, amplitude, period, and phase terms. The stepwise change in refractive index between the silica fiber, hydrogel, and fluid can be modeled as two weak mirrors drawn by the boundary r1 between the fiber and hydrogel, and the boundary r2 between the hydrogel and the surrounding fluid. This system represents a low-finesse Fabry-Perot (FP) cavity. Its model is shown in Figure 6.

[0146] The reflection coefficients are r1 and r2. The second weakest reflection at boundary r2 interferes with the reflected light at boundary r1. This interaction creates a sinusoidal interference pattern in the reflection intensity as a function of wavelength. The FP interference diagram is:

[0147]

number

[0148] It is represented by [this]. During the ceremony,

[0149]

number

[0150] λ is the wavenumber of light inside the hydrogel cavity, and λ is the wavelength of light. Length 39 nm and n g Figure 6 shows the FP reflection spectrum from a hydrogel cavity with a coefficient of 1.35. The absolute length of the hydrogel is determined by finding the period of the cavity (sinus) (resonant optical frequency interval (free spectral range)), and the change in length is determined by finding the phase shift between spectra over time.

[0151] This invention relates to a specific use of a biosensor for measuring glucose concentration. Accordingly, the biosensor of the present invention is suitable for use in in vivo diagnostic methods. The method preferably includes a step of measuring the glucose concentration in the blood of a subject (e.g., a human subject) using the biosensor of the present invention as described above. Therefore, it should be understood that the biosensor of the present invention is suitable for use in a method for determining the glucose concentration in the blood of a subject.

[0152] Preferably, since measurements can be performed continuously or repeatedly at specific time intervals, it is possible to obtain not only a single measurement point that provides limited information about the subject / patient's condition, but also trends in blood glucose concentration over time or a series of measurements. This is particularly important when monitoring a patient's condition over a longer period, for example, when monitoring a patient's postoperative recovery or monitoring such a patient in an intensive care unit. Such repeated or continuous measurements can also be called glucose monitoring. Thus, the present invention also relates to an in vivo glucose monitoring method in a subject, which includes repeatedly measuring the glucose concentration in the subject's blood by using the biosensor of the present invention. Accordingly, the biosensor for measuring glucose concentration of the present invention is provided for use in an in vivo glucose monitoring method.

[0153] Measuring blood glucose levels provides useful information regarding conditions, diseases, and disorders characterized by pathological blood glucose levels (i.e., levels outside the normal range). A blood glucose level below normal, typically below 70 mg / dL (or 3.9 mmol / L), can be called hypoglycemia. A blood glucose level above normal, typically above 200 mg / dL (or 11.1 mmol / L), can be called hyperglycemia. Such conditions can be determined or diagnosed by direct measurement of glucose concentration in the blood. Therefore, the present invention further relates to a method for diagnosing hyperglycemia or hypoglycemia in a subject, the method comprising a procedure for measuring blood glucose levels using the biosensor of the present invention. Accordingly, the present invention provides a biosensor for measuring glucose concentration according to a first embodiment of the present invention for use in in vivo diagnostic methods for hyperglycemia or hypoglycemia. Furthermore, the present invention also relates to using the biosensor of the present invention in an in vitro method for diagnosing hyperglycemia or hypoglycemia, for example, by measuring glucose concentration in a blood sample obtained from a subject.

[0154] Accordingly, the present invention also provides a diagnostic method (particularly an in vitro diagnostic method) that includes a procedure for measuring the blood glucose concentration in a blood sample taken from a subject using the biosensor of the present invention. Thus, the biosensor of the present invention can also be used outside the patient's body to measure the glucose concentration in a sample obtained from the patient. The method may also be called an in vitro diagnostic method. In other words, the present invention provides the biosensor of the present invention for use in an in vitro diagnostic method. The method itself is not particularly limited as long as it includes the necessary procedure for measuring the glucose concentration in a blood sample. The measurement can be carried out, for example, by measuring the volume change of a polymer that occurs in a glucose concentration-dependent manner, as described above.

[0155] The measurement of glucose concentration using the biosensor of the present invention is not limited to measurement in blood. Therefore, the present invention generally relates to the use of the biosensor of the present invention for measuring glucose concentration in samples, including non-blood samples such as urine. However, the sample is preferably a blood sample (e.g., whole blood sample, serum sample, or plasma sample).

[0156] The biosensor of the present invention can be configured to access the target blood through an indwelling arterial catheter. Since the sensor can be applied without interfering with the use of the catheter, the sensor of the present invention does not require a new catheter or the replacement of other devices.

[0157] As further provided herein, the present invention also relates to the use of polymers provided by the present invention for the manufacture of reagents or biosensors for monitoring blood glucose levels in a subject. It should be understood that measurements can be carried out by the changes in the properties of the polymers when they come into contact with various concentrations of glucose.

[0158] As discussed above, the biosensors of the present invention can also be used therapeutically, for example, when incorporated into a glucose concentration-dependent release formulation. Therefore, the present invention further provides a glucose concentration-sensitive release formulation comprising the polymer described herein. The corresponding glucose concentration-sensitive release formulation further comprises one or more pharmaceutically acceptable carriers and an active substance / therapeutic agent (e.g., insulin) delivered in a glucose concentration-dependent manner.

[0159] Such formulations can be manufactured by techniques known in the art, for example, the techniques published in “Remington: The Science and Practice of Pharmacy,” Pharmaceutical Press, 22nd edition. Since release into the bloodstream is preferable when measuring blood glucose levels, the formulations can be formulated as dosage forms for parenteral administration, such as intramuscular, intravenous, subcutaneous, intra-arterial, or intracardiac administration. Dosage forms for parenteral administration include, for example, solutions, emulsions, suspensions, dispersants, and powders and granules for reconstitution. Emulsions are a preferred dosage form for parenteral administration.

[0160] Therefore, when the formulation is administered parenterally, examples of such administration include one or more of the intravenous, intra-arterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intrasternal, intracardiac, intracranial, intramuscular, or subcutaneous administration of the formulation, and / or administration by the use of infusion techniques. In the case of parenteral administration, the formulation containing the polymer of the present invention is best used in the form of a sterile aqueous solution, which may contain other substances such as sufficient salts or sugars to make the solution isotonic with blood. The aqueous solution should be appropriately buffered if necessary (preferably to physiological pH). The manufacture of a suitable parenteral formulation under sterile conditions is readily achieved by standard pharmaceutical techniques.

[0161] The formulations of the present invention can also be formulated as sustained-release systems that may include a semipermeable polymer matrix in the form of molded articles such as films or microcapsules. The sustained-release matrix may be, for example, polylactide, a copolymer of L-glutamic acid and gamma-ethyl-L-glutamate, poly(2-hydroxyethyl methacrylate), ethylene vinyl acetate, or poly-D-(-)-3-hydroxybutyric acid.

[0162] It is preferable that conditions / diseases / disorders related to pathological blood glucose levels be treated. Accordingly, in one embodiment, the present invention relates to a biosensor or glucose concentration-sensitive release formulation for use in the treatment of conditions / diseases / disorders dependent on blood glucose levels. An example of such a condition is diabetes, such as type 1 diabetes. As is well known in the art, diabetes is typically treated by administering insulin to the patient. Accordingly, the formulation of the present invention may further contain insulin.

[0163] However, it is preferable that the biosensor of the present invention is not intended for therapeutic use and / or does not contain insulin. The present invention will be described by the following examples, but these examples are for illustrative purposes only and should not be construed as limiting the scope of the claims. [Examples]

[0164] Name and abbreviation 3APB:3-Acrylamide Phenylboronic Acid 2APB:2-Acrylamide phenylboronic acid ADAM: 5-acrylamide-2-((dimethylamino)methyl)phenylboronic acid TMAPAA: (3-acrylamidopropyl)trimethylammonium chloride PBS: Phosphate-buffered saline GBM: Glucose-binding molecule The structural formulas of the specific compounds described in the experimental section are provided below.

[0165] [ka]

[0166] Materials and methods Chemicals Acrylamide, methylenebisacrylamide, N-hydroxyethylacrylamide (HEAA), 1-hydroxycyclohexylphenyl ketone, 3-(trimethoxysilyl)propyl methacrylate, and squalane were purchased from Sigma-Aldrich. 5-Acrylamide-2-((dimethylamino)methyl)phenylboronic acid (ADAM) and 2-acrylamide phenylboronic acid (2APB) were purchased from Combi-Blocks Inc. 3-Acrylamide phenylboronic acid (3APB), (3-acrylamidepropyl)trimethylammonium chloride (TMAPAA), and N-(3-aminopropyl)methacrylamide hydrochloride (APMA) were purchased from Sigma-Aldrich. N-(3-(dimethylamino)propyl)acrylamide (DMAPAA) was purchased from Tokyo Chemical Industry Co.,Ltd. D-(-)-fructose, D-mannitol, sodium L-lactate, and trisodium citrate dihydrate were purchased from Sigma-Aldrich. Dimethyl sulfoxide was purchased from Sigma-Aldrich.

[0167] D,L-homocysteine ​​thiolactone acrylamide was purchased from Specific Polymers. Ethanolamine was purchased from Sigma-Aldrich. 0.01M PBS pH 7.4 (0.132M NaCl) was prepared in-house, disodium hydrogen phosphate was purchased from Alfa Aesar, monosodium phosphate monohydrate was purchased from Sigma-Aldrich, and sodium chloride was purchased from Sigma-Aldrich.

[0168] All chemicals were used without further purification. Fabrication of a hydrogel sensor In a typical hydrogel sensor formulation, the monomers and molar percentages are as follows: 20% boronic acid acrylamide, 75% acrylamide, and 1-2% methylenebisacrylamide. Exemplary compositions are shown in Table 1.

[0169] Dilute the monomer in either deionized (DI) water, 1 M fructose, or mannitol solution in PBS (pH 7.4, 0.132 M NaCl) to a final total monomer concentration of 1.0 or 1.5 M. Add 1-hydroxycyclohexyl phenyl ketone (photoinitiator) to a concentration of 1.5 mM.

[0170] [Table 1]

[0171] The preparation of the hydrogel sensor follows that described in WO2007 / 104974 (incorporated herein by reference). First, optical glass fibers are peeled and cut to produce a uniform cut surface. Next, the cut surface of the fiber is silane-treated to covalently bond the hydrogel to the glass surface. The silane treatment involves first immersing the fiber cut surface in hydrochloric acid (1.0 M) for 15 minutes, then washing with DI water, and then immersing in ethanol containing 3-(trimethoxysilyl)propyl methacrylate (84 mM) for 10 minutes. Excess 3-(trimethoxysilyl)propyl methacrylate is removed by washing the fiber cut surface under a stream of ethanol. A dome-shaped droplet of the pregel solution is applied to the fiber cut surface using a pipette. During this time, both the cut surface and the pregel are inside a larger squalane oil droplet. This oil serves, firstly, to maintain the stability of the pregel droplet, and secondly, it contains an excess of the dissolved photoinitiator 1-hydroxycyclohexyl phenyl ketone (132 mM). The excess photoinitiator in the oil is necessary to induce polymerization without oxygen-free conditions, and the resulting radicals react first with dissolved oxygen in the oil. The higher concentration of the photoinitiator in the oil compared to the pregel droplets also ensures that the photoinitiator does not significantly leach from the pregel droplets into the oil droplets.

[0172] Polymerization of Pregel occurs by irradiating the Pregel droplet with a 340nm light source positioned directly in front of and inside the Pregel droplet (for 5 minutes). After polymerization, the sensor is briefly washed in pentane to remove oil, and then further washed in DI water with 50% ethanol for 15 minutes to remove unreacted monomers. At this point, the sensor is stored in PBS pH 6.0 until testing.

[0173] Configuration of the sensor experiment In a typical experiment, a test solution is prepared at 37°C using 0.01 M PBS at pH 7.4 containing 132 mM NaCl. The sensor is immersed in the test solution and allowed to equilibrate for at least 10 minutes. Then, the signal from the gel cavity is locked to obtain the initial absolute length of the hydrogel, and the experiment is started. At multiple time points, a portion of the glucose in the PBS is added from a 1 M stock solution that has been left to stand for at least 6 hours to reach mutarotatory equilibrium, while continuously monitoring the change in gel length until the desired concentration is reached. The change in length in response to pH is measured by moving the sensor between two test solutions with constant glucose concentration but different pH levels.

[0174] Example 1 Results and Discussion Monoboronic acid-based glucose sensors often suffer from drawbacks such as lower selectivity for glucose compared to other monosaccharides (e.g., fructose) and significant changes in glucose binding with small pH fluctuations. Here, we developed a sensor that exhibits better selectivity for glucose than a range of common interfering substances while minimizing the effects of pH changes.

[0175] One of the most commonly used monoboronic acids for glucose sensing is 3-acrylamide phenylboronic acid (3APB), which is incorporated into materials such as polymers to provide glucose responsiveness. Here, 3APB is used as a reference example.

[0176] The boron center of 3APB has a pKa of approximately 8.8. 3 At physiological pH, it mainly exists in a trigonal sp2 form, which is not suitable for glucose binding. 4,5As a result of the relatively high pKa, even small changes in pH from 7.4 to higher or lower levels significantly increase or decrease the degree of glucose binding, respectively. This has a significant impact on the sensor reading and necessitates complex pH correction. 6,7 Due to this pH sensitivity, using 3APB for in vivo glucose sensing is inappropriate because, in extreme cases, blood pH can range from as low as 6.9 to as high as 7.6. 3APB and positively charged comonomers (as described in various literatures) 7,8,9,10 When the glucose response of a sensor with ) was tested at pH 6.9, 7.4, and 7.6 (Figure 2A), a large deviation was observed between the sensor reading and the reference glucose reading. To address pH interference, a Wulff-type sensor was used. 2 Boronic acid 5-acrylamido-2-((dimethylamino)methyl)phenylboronic acid (ADAM) was selected as GBM and replaced with 3APB. Because Wulff-type boronic acid has an amino group in the ortho position relative to boronic acid, the pKa of boronic acid is reduced to approximately 5.2. 2,11 It is suitable for glucose binding at physiological pH. Wulff-type boronic acid has been successfully used in fluorescent glucose sensors. 12,13 The precise mechanism of the interaction between the amino group and boron is still under discussion. 14,15 .

[0177] Despite the lower pKa, Wulff-type boronic acid, compared to other monoboronic acid, likely due to steric hindrance. 5 It has been reported to have low affinity for glucose. 11,16 In fact, sensors constructed using ADAM required 2.5 to 3.5 times more mol% of GBM than those using 3APB to obtain a similar glucose response. Nevertheless, with proper optimization of the ADAM concentration, a large, linear change (contraction) in the length of the ADAM hydrogel sensor was achieved, suitable for glucose detection within the physiological range (Figure 3A). The ADAM-based sensor, in contrast to the 3APB sensor, 9 Furthermore, no additional cationic monomers were needed to obtain a linear contraction induced by glucose (Figure 3).

[0178] Importantly, the ADAM sensor exhibits significantly lower pH dependence compared to the 3APB sensor (Figure 2), indicating that the ADAM sensor is suitable for in vivo glucose monitoring.

[0179] The ADAM sensor's superior selectivity for glucose compared to other common interfering substances was found to be enhanced by the addition of the quaternary ammonium comonomer TMAPAA (Figure 3). Other groups have also reported similar effects when using cationic comonomers with 3APB, indicating that a positive charge is necessary to achieve an acceptable glucose response with 3APB at physiological pH. 17,18,10 .

[0180] Compared to other examples, a considerably smaller ratio of cationic monomer to boronic acid was necessary to almost completely suppress interference from fructose (Figure 2B). 17,18,10 .

[0181] The addition of TMAPAA did not significantly affect the glucose-mediated sensor response (Figure 3A). Importantly, a low mol% concentration of cationic groups in the hydrogel was essential to avoid unwanted interference with salts (Figure 3D). Sensors with 3% TMAPAA and 20% ADAM showed negligible interference with fructose, mannitol, lactate, and citrate, even though the concentrations of the tested interfering substances far exceeded those expected in vivo. Furthermore, they exhibited reduced interference with a pH change from 6.9 to 7.6 (Figures 3 and 4), demonstrating selectivity not typically observed in monoboronic acids. This observed selectivity is thought to be partly driven by the ADAM hydrogel's ability to form a 2:1 complex with glucofuranose. 1 .

[0182] 2APB, recently studied as GBM, offers significantly lower pH interference than 3APB due to internal coordination between boron and adjacent carbonyl groups. 19,20,21The sensor constructed using 2APB showed a slow reaction rate with glucose, resulting in only slight contraction at 6 and 20 mM glucose, but with very little interference from lactate, citrate, and pH changes. Unexpectedly, the 2APB sensor showed a large and rapid contraction at 5 mM mannitol (Figure 4). The mannitol response of the 2APB sensor was significantly larger for glucose or fructose compared to ADAM or 3APB sensors (Table 2). The reason for this response is unknown, but it may be due to crosslinking between 2APB and mannitol. 22 Considering that the 2APB sensor contracts with mannitol and the ADAM sensor expands with mannitol, it was hypothesized that adding 2APB to the ADAM sensor at a specific mol% might selectively suppress mannitol interference. By changing the molar ratio of ADAM and 2APB, it was found that mannitol interference at 2.2 mM glucose could be suppressed when using 3% 2APB and 25% ADAM (Figure 5).

[0183] [Table 2]

[0184] Example 2 In this example, the sensitivity of sensors containing the polymer hydrogel described below to glucose was tested according to the method outlined above, and the results are summarized in Table 3.1 below.

[0185] [Table 3-1]

[0186] The results are shown in Figure 7A. For the same monomer ratio, the change in length of the ADAM-acrylamide sensor at 20 mM glucose is approximately 1.6 times greater than that of the ADAM-HEAA sensor. Therefore, the acrylamide sensor is more sensitive to glucose. It is noteworthy that in both cases (acrylamide and HEAA), the sensors contract in response to glucose.

[0187] The experiment was repeated with polymer hydrogels that did not contain TMAPAA. These were obtained according to the method outlined above and are summarized in Table 3.2 below.

[0188] [Table 3-2]

[0189] The results are shown in Figure 7B. It can be concluded that, at the same monomer ratio, the change in length of the ADAM-acrylamide sensor at 20 mM glucose is approximately twice as large as that of the ADAM-HEAA sensor. Therefore, the acrylamide sensor is more sensitive to glucose. Notably, in both cases, the (acrylamide and HEAA) sensors contract in response to glucose.

[0190] Example 3 Three different cationic monomers were introduced into polymer gel formulations, and the resulting biosensors were tested for their ability to withstand interference from the presence of citrate. The monomers introduced are listed below.

[0191] [ka]

[0192] The properties of the obtained hydrogels are shown in Table 4 below. The total monomer concentration in all formulations is 1.5 M.

[0193] [Table 4]

[0194] The results of expansion measurements performed with these hydrogels are summarized in Figure 8. Sensors without cationic comonomers expanded significantly with citrate. This indicates citrate interference, which is undesirable for clinically relevant measurements. The cationic monomers TMAPAA, DMAPAA, and APMA were found to reduce citrate interference without introducing a significant change in the sensor's response to glucose.

[0195] Example 4 A polymer hydrogel containing the ADAM moiety was manufactured according to a polymer hydrogel manufacturing method, and then modified to obtain the polymer hydrogel of the present invention. 23 .

[0196] A monomer solution containing 16 or 28 mol% thiolactone acrylamide was photopolymerized to form the corresponding hydrogel. The hydrogel sensor was then incubated with 26 mM alkene (ADAM) in a 1:1 DMSO-PBS solution. After a short time, ethanolamine was added (final concentration 5 M) to initiate the reaction. The reaction was left overnight at room temperature with stirring, after which the sensor was washed with 50% ethanol in DI water for 15 minutes and then tested.

[0197] [ka]

[0198] The properties of the obtained hydrogel are shown in Table 5 below.

[0199] [Table 5]

[0200] The results are summarized in Figure 9. The ADAM sensor can also be constructed by thiol-ene type post-polymerization modification. As shown in Figure 9, greater contraction by glucose is observed in the post-modified sensor (Figure 9A) compared to the directly polymerized sensor (Figure 9B). However, it should be reiterated that very similar behavior is observed in both types of sensors.

[0201] References 1. Eggert, H., Frederiksen, J., Morin, C. & Norrild, JC A new glucose-selective fluorescent bisboronic acid. First report of strong α-furanose complexation in aqueous solution at physiological pH. J. Org. Chem. 64, 3846-3852 (1999). 2. Wulff, G. Selective binding to polymers via covalent bonds. the construction of chiral cavities as specific receptor sites. Pure Appl. Chem. 54, 2093-2102 (1982). 3. Springsteen, G. & Wang, B. A detailed examination of boronic acid-diol complexation. Tetrahedron58, 5291-5300 (2002). 4. Bosch, LI, Fyles, TM & James, TD Binary and ternary phenylboronic acid complexes with saccharides and Lewis bases. Tetrahedron 60, 11175-11190 (2004). 5. Yan, J., Springsteen, G., Deeter, S. & Wang, B. The relationship among pK a, pH, and binding constants in the interactions between boronic acids and diols - It is not as simple as it appears. Tetrahedron 60, 11205-11209 (2004). 6. Worsley, G. J. et al. Measurement of glucose in blood with a phenylboronic acid optical sensor. J. Diabetes Sci. Technol. 2, 213-220 (2008). 7. Tierney, S., Hasle Falch, B. M., Hjelme, D. R. & Stokke, B. T. Determination of glucose levels using a functionalized hydrogel-optical fiber biosensor: Toward continuous monitoring of blood glucose in vivo. Anal. Chem. 81, 3630-3636 (2009). 8. Horkay, F. et al. Thermodynamic analysis of the selectivity enhancement obtained by using smart hydrogels that are zwitterionic when detecting glucose with boronic acid moieties. Sensors Actuators, B Chem. 160, 1363-1371 (2011). 9. Lin, G. et al. Osmotic swelling pressure response of smart hydrogels suitable for chronically implantable glucose sensors. Sensors Actuators, B Chem. 144, 332-336 (2010). 10. Horgan, A. M. et al. Crosslinking of phenylboronic acid receptors as a means of glucose selective holographic detection. Biosens. Bioelectron. 21, 1838-1845 (2006). 11. Brooks, W. L. A., Deng, C. C. & Sumerlin, B. S. Structure-Reactivity Relationships in Boronic Acid-Diol Complexation. ACS Omega 3, 17863-17870 (2018). 12. Crane, B. C. et al. The development of a continuous intravascular glucose monitoring sensor. J. Diabetes Sci. Technol. 9, 751-761 (2015). 13. Mortellaro, M. & DeHennis, A. Performance characterization of an abiotic and fluorescent-based continuous glucose monitoring system in patients with type 1 diabetes. Biosens. Bioelectron. 61, 227-231 (2014). 14. Sun, X. et al. The mechanisms of boronate ester formation and fluorescent turn-on in ortho-aminomethylphenylboronic acids. Nat. Chem. 11, 768-778 (2019). 15. Ortega-Valdovinos, L. R. & Yatsimirsky, A. K. Probing the Role of the Bridging Nitrogen in the Signaling Mechanism of an Anthracene-Boronic Acid Sugar Sensor and a Different Version of the PET-Based Mechanism. J. Org. Chem. (2023) doi:10.1021 / acs.joc.3c00129. 16. Dowlut, M. & Hall, D. G. An improved class of sugar-binding boronic acids, soluble and capable of complexing glycosides in neutral water. J. Am. Chem. Soc.128, 4226-4227 (2006). 17. Nguyen, T., Magda, J. J. & Tathireddy, P. Manipulation of the isoelectric point of polyampholytic smart hydrogels in order to increase the range and selectivity of continuous glucose sensors. Sensors Actuators, B Chem. 255, 1057-1063 (2018). 18. Tierney, S., Volden, S. & Stokke, B. T. Glucose sensors based on a responsive gel incorporated as a Fabry-Perot cavity on a fiber-optic readout platform. Biosens. Bioelectron. 24, 2034-2039 (2009). 19. Tang, Z., Guan, Y. & Zhang, Y. The synthesis of a contraction-type glucose-sensitive microgel working at physiological temperature guided by a new glucose-sensing mechanism. Polym. Chem. 9, 1012-1021 (2018). 20. Zhang, C., Losego, M. D. & Braun, P. V. Hydrogel-based glucose sensors: Effects of phenylboronic acid chemical structure on response. Chem. Mater. 25, 3239-3250 (2013). 21. Yang, X., Pan, X., Blyth, J. & Lowe, C. R. Towards the real-time monitoring of glucose in tear fluid: Holographic glucose sensors with reduced interference from lactate and pH. Biosens. Bioelectron. 23, 899-905 (2008). 22. Lopalco, A., Marinaro, W. A., Day, V. W. & Stella, V. J. Isolation, Solubility, and Characterization of D-Mannitol Esters of 4-Methoxybenzeneboronic Acid. J. Pharm. Sci. 106, 601-610 (2017). 23. Espeel, P., Goethals, F., Stamenovic, M. M., Petton, L. & Du Prez, F. E. Double modular modification of thiolactone-containing polymers: Towards polythiols and derived structures. Polym. Chem. 3, 1007-1015 (2012).

Claims

1. A biosensor for measuring glucose concentration, wherein the biosensor is defined by formula (I): 【Chemistry 1】 A polymer containing a portion thereof, Here, the portion of formula (I) is fixed to the polymer, In the above formula, each R is independently C １－５ Alkyl, C ２－５ Alkenyl or C ２－５ It is alkinyl, In the above formula, each R Ｓ is independently C １－５ alkyl, C ２－５ alkenyl, C ２－５ alkynyl, -(C ０－３ alkylene)-OH, -(C ０－３ alkylene)-O(C １－５ alkyl), -(C ０－３ alkylene)-O(C １－５ alkylene)-OH, -(C ０－３ alkylene)-O(C １－５ alkylene)-O(C １－５ alkyl), -(C ０－３ alkylene)-SH, -(C ０－３ alkylene)-S(C １－５ alkyl), -(C ０－３ alkylene)-S(C １－５ alkylene)-SH, -(C ０－３ alkylene)-S(C １－５ alkylene)-S(C １－５ alkyl), -(C ０－３ alkylene)-NH ２ , -(C ０－３ alkylene)-NH(C １－５ alkyl), -(C ０－３ alkylene)-N(C １－５ alkyl)(C １－５ alkyl), -(C ０－３ alkylene)-NH-OH, -(C ０－３ alkylene)-N(C １－５ alkyl)-OH, -(C ０－３ alkylene)-NH-O(C １－５ alkyl), -(C ０－３ alkylene)-N(C １－５ alkyl)-O(C １－５ alkyl), -(C ０－３ alkylene)-halogen, -(C ０－３ alkylene)-(C １－５ haloalkyl), -(C ０－３ alkylene)-O-(C １－５ haloalkyl), -(C ０－３ alkylene)-CN, -(C ０－３ alkylene)-NO ２ , -(C ０－３ -(alkylene)-CHO, -(C ０－３ -(alkylene)-CO-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-COOH, -(C ０－３ -(alkylene)-CO-O-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-O-CO-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-CO-NH ２ , -(C ０－３ -(alkylene)-CO-NH(C １－５ -(alkyl), -(C ０－３ -(alkylene)-CO-N(C １－５ -(alkyl)(C １－５ -(alkyl), -(C ０－３ -(alkylene)-NH-CO-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-N(C １－５ -(alkyl)-CO-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-NH-CO-O-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-N(C １－５ -(alkyl)-CO-O-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-O-CO-NH-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-O-CO-N(C １－５ -(alkyl)-(C １－５ -(alkyl), -(C ０－３ -(alkylene)-SO ２ -NH ２ , -(C ０－３ -(alkylene)-SO ２ -NH(C １－５ -(alkyl), -(C ０－３ -(alkylene)-SO ２ -N(C １－５ -(alkyl)(C １－５ -(alkyl), -(C ０－３ -(alkylene)-NH-SO ２ -(C １－５ -(alkyl), -(C ０－３ - (alkylene)-N(C １－５ alkyl)-SO ２ -(C １－５ alkyl), -(C ０－３ alkylene)-SO ２ -(C １－５ alkyl), -(C ０－３ alkylene)-SO-(C １－５ alkyl), -(C ０－３ alkylene)-carbocyclic, and -(C ０－３ alkylene)-heterocyclic, wherein the carbocyclic moiety of the -(C ０－３ alkylene)-carbocyclic and the heterocyclic moiety of the -(C ０－３ alkylene)-heterocyclic are each independently selected from C １－４ alkyl, halogen, -CN, -NO ２ , -OH, -O-(C １－４ alkyl), -SH, -S-(C １－４ alkyl), -NH ２ , -NH(C １－４ alkyl), -N(C １－４ alkyl)(C １－４ alkyl), -COOH, -COO(C １－４ alkyl), -CONH ２ , -CONH(C １－４ alkyl), -CON(C １－４ alkyl)(C １－４ alkyl), -NHCO(C １－４ alkyl) and -N(C １－４ alkyl)-CO(C １－４ alkyl) and may be substituted with one or more groups independently selected therefrom; In the above formula, n is 0, 1, 2, or 3. Biosensor.

2. The biosensor according to claim 1, wherein each R is methyl.

3. The biosensor according to claim 1 or 2, wherein n is 0 or 1, preferably n is 0.

4. The polymer is given by formula (Ia): 【Chemistry 2】 The formula includes the part, and in the formula, R, R Ｓ The biosensor according to any one of claims 1 to 3, wherein n is as described in any one of claims 1 to 3.

5. The biosensor according to any one of claims 1 to 4, wherein the polymer is a polymeric hydrogel.

6. The biosensor according to any one of claims 1 to 5, wherein the polymer is an unsubstituted polyacrylamide-based polymer.

7. The biosensor according to any one of claims 1 to 6, wherein the polymer further comprises a portion that is positively charged at pH = 7.

4.

8. The polymer further, -NH ２ , -NH ３ ＋ ,-NH(C １－５ Alkyl), - (NH ２ (C １－５ Alkyl)) ＋ , -N(C １－５ Alkyl) ２ ,-(NH(C) １－５ Alkyl) ２ ) ＋ , and -N(C) １－５ Alkyl) ３ ＋ A biosensor according to any one of claims 1 to 6, comprising a portion containing a group selected from.

9. The polymer further forms formula (II): 【Transformation 3】 A biosensor according to any one of claims 1 to 6, including the portion.

10. The polymer further forms formula (III): 【Chemistry 4】 A biosensor according to any one of claims 1 to 9, including the portion.

11. polymers (i) Can be obtained by polymerization reaction of a composition comprising an acrylamide monomer containing a boronic acid-based glucose-bonding moiety of formula (I), Preferably (i) 【Transformation 5】 [In the formula, R, R Ｓ The biosensor according to any one of claims 1 to 10, wherein n is as described in any one of claims 1 to 10.

12. The composition used in the polymerization reaction is further, (ii) Acrylamide monomers that do not contain the boronic acid portion, and (iii) Acrylamide monomer as a crosslinking agent Includes, Preferably (ii) 【Transformation 6】 It comprises a monomer selected from [wherein q is an integer from 1 to 10]; and / or Equation (iii) 【Transformation 7】 The biosensor according to claim 11, comprising a monomer selected from [wherein q is an integer from 1 to 10].

13. The biosensor according to any one of claims 1 to 12, wherein the polymer contracts in the presence of glucose, preferably the rate of contraction is substantially linearly proportional to the glucose concentration.

14. The biosensor according to claim 1, wherein each R is methyl, n is 0, and the polymer is an unsubstituted polyacrylamide-based polymer.

15. The biosensor according to claim 1, wherein each R is methyl, n is 0, and the polymer further includes a portion that is positively charged at pH = 7.

4.

16. Each R is methyl, n is 0, and the polymer is further -NH ２ , -NH ３ ＋ ,-NH(C １－５ Alkyl), -(NH ２ (C １－５ Alkyl)) ＋ , -N(C １－５ Alkyl) ２ ,-(NH(C) １－５ Alkyl) ２ ) ＋ , and -N(C) １－５ Alkyl) ３ ＋ The biosensor according to claim 1, comprising a portion containing a group selected from.

17. The biosensor according to any one of claims 1 to 16, wherein the glucose-dependent contraction rate is substantially pH-independent, preferably in the range of 6.9 to 7.6, and preferably in the range of 7.4 to 7.

6.

18. A biosensor according to any one of claims 1 to 17, for use in diagnosis.

19. A biosensor according to any one of claims 1 to 17, for use in monitoring blood glucose levels.

20. The biosensor according to claim 19, wherein blood glucose monitoring is performed on a subject in an intensive care unit, and / or blood glucose monitoring is performed on an unconscious subject.

21. A polymer as defined in any one of claims 1 to 16.

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